Humanized CC chemokine receptor 4 (CCR4) antibodies and methods of use thereof
The present invention provides humanized monoclonal antibodies, bi-specific antibodies, antibody conjugates, and fusion proteins that bind to the chemokine receptor CCR4. This antibody is derived from CCR4-IgG1 and recognizes the same epitope. This antibody contains either an IgG4 or a stabilized IgG4 in order to improve binding efficiency and reduce in vivo Fab arm exchange. Binding of the antibodies disclosed herein to CCR4 inhibits ligand-mediated activities and is used to treat symptoms of cancer.
1. An isolated humanized monoclonal antibody that binds to the human CC chemokine receptor 4 (CCR4) and has an IgG4 heavy chain constant region, wherein the antibody comprises:
a. a heavy chain with three CDRs comprising a CDR1 comprising amino acid sequence SEQ ID NO: 9, a CDR 2 comprising amino acid sequence SEQ ID NO:11, and a CDR 3 comprising amino acid sequence SEQ ID NO: 13; and
a light chain with three CDRs comprising a CDR1 comprising amino acid sequence SEQ ID NO: 10, a CDR 2 comprising amino acid sequence SEQ ID NO: 12, and a CDR 3 comprising amino acid sequence SEQ ID NO: 14;
b. a heavy chain with three CDRs comprising a CDR1 comprising amino acid sequence SEQ ID NO: 32, a CDR 2 comprising amino acid sequence SEQ ID NO: 11, and a CDR 3 comprising amino acid sequence SEQ ID NO: 34; and
a light chain with three CDRs comprising a CDR1 comprising amino acid sequence SEQ ID NO: 10, a CDR 2 comprising amino acid sequence SEQ ID NO: 12, and a CDR 3 comprising amino acid sequence SEQ ID NO: 39;
c. a heavy chain with three CDRs comprising a CDR1 comprising amino acid sequence SEQ ID NO: 33, a CDR 2 comprising amino acid sequence SEQ ID NO: 11, and a CDR 3 comprising amino acid sequence SEQ ID NO: 35; and
a light chain with three CDRs comprising a CDR1 comprising amino acid sequence SEQ ID NO: 10, a CDR 2 comprising amino acid sequence SEQ ID NO: 12, and a CDR 3 comprising amino acid sequence SEQ ID NO: 40;
d. a heavy chain with three CDRs comprising a CDR1 comprising amino acid sequence SEQ ID NO: 33, a CDR 2 comprising amino acid sequence SEQ ID NO: 11, and a CDR 3 comprising amino acid sequence SEQ ID NO: 36; and
a light chain with three CDRs comprising a CDR1 comprising amino acid sequence SEQ ID NO: 10, a CDR 2 comprising amino acid sequence SEQ ID NO: 12, and a CDR 3 comprising amino acid sequence SEQ ID NO: 41;
e. a heavy chain with three CDRs comprising a CDR1 comprising amino acid sequence SEQ ID NO: 33, a CDR 2 comprising amino acid sequence SEQ ID NO: 11, and a CDR 3 comprising amino acid sequence SEQ ID NO: 37; and
a light chain with three CDRs comprising a CDR1 comprising amino acid sequence SEQ ID NO: 10, a CDR 2 comprising amino acid sequence SEQ ID NO: 12, and a CDR 3 comprising amino acid sequence SEQ ID NO: 42; or
f. a heavy chain with three CDRs comprising a CDR1 comprising amino acid sequence SEQ ID NO: 31, a CDR 2 comprising amino acid sequence SEQ ID NO: 11, and a CDR 3 comprising amino acid sequence SEQ ID NO: 13; and
a light chain with three CDRs comprising a CDR1 comprising amino acid sequence SEQ ID NO: 10, a CDR 2 comprising amino acid sequence SEQ ID NO: 12, and a CDR 3 comprising amino acid sequence SEQ ID NO: 38; and
a heavy chain constant region comprising the amino acid sequence of SEQ ID NO: 6 or SEQ ID: 8, and wherein the antibody modulates regulatory T cell recruitment without inducing T lymphocyte depletion.
2. The antibody of claim 1 , wherein the antibody comprises:
a. a variable heavy chain region (VH) comprising the amino acid sequence of SEQ ID NO: 2 and a variable light chain region (VL) comprising the amino acid sequence of SEQ ID NO: 4;
b. a variable heavy chain region (VH) comprising the amino acid sequence of SEQ ID NO: 16 and a variable light chain region (VL) comprising the amino acid sequence of SEQ ID NO: 18;
c. a variable heavy chain region (VH) comprising the amino acid sequence of SEQ ID NO: 20 and a variable light chain region (VL) comprising the amino acid sequence of SEQ ID NO: 22;
d. a variable heavy chain region (VH) comprising the amino acid sequence of SEQ ID NO: 24 and a variable light chain region (VL) comprising the amino acid sequence of SEQ ID NO: 26;
e. a variable heavy chain region (VH) comprising the amino acid sequence of SEQ ID NO: 28 and a variable light chain region (VL) comprising the amino acid sequence of SEQ ID NO: 30; or
f. a variable heavy chain region (VH) comprising the amino acid sequence of SEQ ID NO: 44 and a variable light chain region (VL) comprising the amino acid sequence of SEQ ID NO: 46.
3. The antibody of claim 2 , wherein said antibody has a binding affinity of about 1. 5 nm −1 or less.
4. The antibody according to claim 1 , wherein the antibody is linked to a therapeutic agent.
5. The antibody of claim 4 , wherein said therapeutic agent is a toxin, a radiolabel, a siRNA, a small molecule, or a cytokine.
6. The antibody of claim 5 wherein said cytokine is IL-2 or TGF-beta.
7. A bi-specific antibody comprising the antibody of claim 1 and an antibody that immunospecifically binds to a second antigen.
8. The bi-specific antibody of claim 7 , wherein the second antigen is a tumor associated antigen or a T-cell function modulating molecule.
9. The bi-specific antibody of claim 8 , wherein the tumor associated antigen is CA-IX, ErbB2 or HVEM.
10. A cell producing the antibody of claim 1 .
11. A method of inhibiting the migration of regulatory T-cells (Tregs) in a subject by administering to said subject the antibody of claim 1 .
12. A method of claim 11 , wherein lymphocytes are not depleted.
13. A method of claim 11 , wherein effector T-cells are not depleted.
14. A method of claim 11 , wherein Tregs are not depleted.
15. A method of augmenting an immune response to an antigen in a subject comprising administering to the subject the antibody of claim 1 .
16. The method of claim 15 , wherein said antigen is a viral antigen, a bacterial antigen or a tumor associated antigen.
17. The method of claim 15 , wherein said administration of said antibody causes an increase in antigen specific T-cell activity.
18. The method of claim 15 , wherein said administration of said antibody causes an increase in T-cell proliferation.
19. The method of claim 15 , wherein effector T-cells are augmented.
20. A method of reversing regulatory T cell-mediated suppression of effector T cell proliferation comprising contacting a T cell with the antibody of claim 1 .
21. A method of treating or alleviating a symptom of cancer, comprising administering to a subject in need thereof a composition comprising an antibody according to claim 1 .
22. The method of claim 21 , wherein said cancer is a solid cancer or a hematologic cancer.
23. The method of claim 22 , wherein said hematologic cancer is cutaneous T-cell Lymphoma (CTCL), mycosis fungoides (MF), primary cutaneous anaplastic large cell Lymphoma (cutaneous ALCL), Sezary syndrome, or adult T cell Leukemia/Lymphoma (ATLL).
24. The method of claim 22 , wherein the cancer is a solid cancer or a cancer that overexpresses CA IX, PD-LI, or HVEM.
25. The method of claim 22 , were said solid cancer is renal cell carcinoma, breast cancer, lung cancer, ovarian cancer, prostate cancer, colon cancer, cervical cancer, brain cancer, liver cancer, pancreatic cancer, kidney or stomach cancer.